The regulation of adenovirus gene expression is being studied by a combined biochemical and genetic approach. Existing temperature-sensitive mutants of adenovirus type 5 are being used to define the relationship between virus DNA replication and late virus transcription. The mechanism and specificity of the shut-off of early transcription by the virus 72K protein is being studied by comparing the kinetics of virus transcription in isolated nuclei from cells infected by wt adenovirus and by ts125. Specificity of the adenovirus 72K protein effect on transcription is also being studied in monkey cells coinfected by adenovirus and wt SV40 or tsA mutants. New temperature-sensitive adenovirus mutants will be sought by using selective techniques to enrich for mutants defective in late gene expression. These mutants will be characterized for ability to replicate virus DNA and express late genes. Revertants of ts125 are being screened for overproduction of the 72K DNA-binding protein, and for aberrant effects on early virus transcription. The ability of these revertants to transform rodent cells in culture will be tested.